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1 Department of Physiology, Dartmouth Medical School, Hanover, New Hampshire 03755; and 2 Molecular Medicine and Renal Units, Beth Israel Deaconess Medical Center, and Departments of Medicine and Cell Biology, Harvard Medical School, Boston, Massachusetts 02215
Serous cells secrete
Cl
and HCO3 and play an important
role in airway function. Recent studies suggest that a
Cl/HCO3 anion exchanger (AE) may
contribute to Cl secretion by airway epithelial cells.
However, the molecular identity, the cellular location, and the
contribution of AEs to Cl secretion in serous epithelial
cells in tracheal submucosal glands are unknown. The goal of the
present study was to determine the molecular identity, the cellular
location, and the role of AEs in the function of serous epithelial
cells. To this end, Calu-3 cells, a human airway cell line with a
serous-cell phenotype, were studied by RT-PCR, immunoblot, and
electrophysiological analysis to examine the role of AEs in
Cl secretion. In addition, the subcellular location of AE
proteins was examined by immunofluorescence microscopy. Calu-3 cells
expressed mRNA and protein for AE2 as determined by RT-PCR and Western
blot analysis, respectively. Immunofluorescence microscopy identified AE2 in the basolateral membrane of Calu-3 cells in culture and rat
tracheal serous cells in situ. In
Cl/HCO3/Na+-containing
media, the 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate
(CPT-cAMP)-stimulated short-circuit anion current (Isc) was reduced by basolateral but not by
apical application of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid
(50 µM) and 4,4'-dinitrostilbene-2,2'-disulfonic acid [DNDS (500 µM)], inhibitors of AEs. In the absence of Na+ in the
bath solutions, to eliminate the contributions of the Na+/HCO3 and
Na+/K+/2Cl cotransporters to
Isc, CPT-cAMP stimulated a small DNDS-sensitive Isc. Taken together with previous studies, these
observations suggest that a small component of cAMP-stimulated
Isc across serous cells may be referable to
Cl secretion and that uptake of Cl across
the basolateral membrane may be mediated by AE2.
cystic fibrosis; anion exchanger 2; serous cell; submucosal gland; chloride transport; sodium bicarbonate cotransport
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